In the United States, herpes simplex virus type 1 (HSV-1) is the most common cause of corneal blindness due to an infectious agent. Most ocular herpes disease is due to viral reactivations from latently infected neurons of the trigeminal ganglia (TG), rather than to primary acute infection. Prevention of reactivations requires a complex interplay between the virus, the neurons, and the immune system. While the host's CD8+ T cells play a critical role in controlling HSV-1 reactivation from latently infected neurons of TG, the virus has evolved immune evasion strategies as a counter-defense to protect itself from being attacked by the host's CD8+ T cells. Understanding the mechanisms of HSV-1 immune evasion during latency has been enhanced by our recent discovery of dysfunctional HSV-specific CD8+ T cells in latently infected TG. This is triggered by the HSV-1 Latency Associated Transcript (LAT), the only viral gene abundantly transcribed during latent infection. Building on strong published and preliminary data, this application focuses on: (1) Understanding the LAT- mediated mechanisms that lead to dysfunctional CD8+ T cells in latently infected TG; and (2) Using that knowledge to devise novel powerful T-cell based immunotherapeutic strategies to prevent reactivations, recurrent eye disease and blindness. We have recently found that: (i) Three T-cell co-inhibitory receptors, PD- 1, TIM-3, and BTLA are highly expressed on HSV-specific CD8+ T cells, but not on HSV-specific CD4+ T cells, in TG from mice latently infected with LAT(+) compared to LAT(-) virus. (ii) A significant proportion of HSV- specific CD8+ T cells in LAT(+) TG are partially dysfunctional. (iii) PD-L1 and HVEM expression are increased in LAT(+) TG compared to LAT(-) TG. These intriguing results, together with related reports by others in the field, lead us to hypothesize that: (1) In HSV-1 latently infected TG, LAT promotes dysfunction of CD8+ T-cells and impairs their immuno-surveillance function through one or several T-cell inhibitory pathways, thus promoting virus reactivation from latently infected neurons; and (2) Reversing the dysfunction of CD8+ T-cells will decrease HSV-1 reactivation, recurrent eye disease, and prevent loss of sight. Our specific Aims are: Aim 1: Determine the kinetics of expression of T-cell co-inhibitory receptors, PD-1, TIM-3, and BTLA, on TG-resident CD8+ T cells of humanized HLA Tg mice ocularly infected with HSV-1 LAT(+) vs. HSV-1 LAT(-) and confirm the role of these co-inhibitory pathways in the dysfunction of CD8+ T-cells seen in latently infected TG. Aim 2: Test the hypothesis that in vivo blocking of PD-1/PD-L1, TIM-3/GAL-9 and/or BTLA/HVEM T-cell inhibitory signaling pathways will restore CD8+ T cell protective function and enhance the efficacy of therapeutic immunization in humanized HLA Tg mice. This translational research, that gathers a multidisciplinary team, addresses the current NEI Audacious Goals Initiative: Development of new treatments through small molecules approach to treat eye disease and to restore sight. Successful completion of this pre-clinical study in humanized HLA Tg mice should lead to a novel and powerful immunotherapy to prevent blinding recurrent herpetic disease.